The advent of DNA-sequence-based approaches to analyze microbial environments has led to a deepened appreciation for the diversity, ubiquity, and functions of microbial life. For instance, the gastrointestinal tract of humans and other vertebrates is colonized by complex microbial communities that promote gut development, nutrient metabolism, and immune homeostasis. Of particular importance to human health, gut microbes have emerged as major risk determinants for obesity and metabolic disorders, in part because of their role in modulating accessibility and absorption of energy-rich dietary nutrients in vertebrates. For example, colonization of germ-free zebrafish with Exiguobacterium sp. ZWU0009, a Firmicutes bacterium originally isolated from the zebrafish intestine, enhanced the ability of intestinal enterocytes to absorb dietary fat. Unfortunately, the molecular bases for how bacteria like Exiguobacterium sp. ZWU0009 colonize the intestine and influence host physiology are poorly understood.
Indeed, most microbes are not amenable to genetic manipulation because methods for robust DNA transformation, insertional mutagenesis, and trans-expression of genes are largely lacking. For a select group of microbial species, including members of the Bacteroides genus, some strains are amenable to transposon mutagenesis and have been invaluable in helping decipher the requirement of individual genes in gut colonization and nutrient homeostasis. However, genetic tools do not exist for the vast majority of intestinal microbes. As a result, the function of individual genes and their contribution to host-microbe and microbe-microbe interactions within the gut often relies on information inferred from homology to genes characterized in phylogenetically unrelated, but genetically tractable, bacterial systems. This reliance on previously characterized genes is a significant challenge that inhibits the functional annotation of novel genes emerging from metagenomic studies, as well as impairs the ability to identify new targets for the development of therapeutic compounds and treatments.
Given the paucity of experimental tools to manipulate bacterial genomes, there is a need for improved methods for determining how microbial communities assemble and influence human and environmental health. Embodiments of the present disclosure address these and other needs.